Hepatitis C virus (HCV) is a major cause of post-transfusion and sporadic hepatitis, leading to chronic liver disease in at least 50% of infected individuals. The host and/or viral factors which permit the establishment of persistent HCV infection in a high percentage of infected individuals are unknown. In many viral infections, cytotoxic T lymphocytes (CTL) are an important component of host immunity, although they may also mediate tissue damage. The role of HCV-specific CTL role in disease protection or pathogenesis is unknown. The goal of this proposal is the identification and characterization of HCV-specific CTL from the liver-infiltrating lymphocytes and peripheral blood mononuclear cells of persons with a variety of clinical manifestations of HCV infection. Recombinant HCV- vaccinia viruses will be used to express HCV structural and non-structural proteins in target cells. CTL will be identified using both specific and non-specific stimuli. The specificities and magnitude of the CTL response will be correlated with the clinical course of the infection. The patterns of cytokine release by virus-specific CTL upon recognition of target cells will be studied using ELISA. In addition, the effect of viral heterogeneity on CTL recognition of targets will be studied using polymerase chain reaction (PCR) to amplify viral sequences containing CTL epitopes. The specific aims of this proposal are: 1) identification of HCV-specific CTL which recognize structural and non-structural viral proteins, and characterization of the spectrum of responses in infected individuals; 2), correlation of the spectrum of HCV-specific CTL responses with disease activity in acutely and chronically infected subjects; 3), characterization of the role of cytokines in HCV-specific CTL responses; and 4), determination of the role of HCV antigenic variation in viral escape from recognition by HCV-specific CTL as a means of viral persistence. The information provided by these studies should allow a comprehensive evaluation of the role of cellular immunity in HCV infection, and should facilitate rational vaccine design and development of new therapeutic agents. During preliminary studies conducted in the past two years, our laboratory has identified HCV-specific CTL within the livers of chronically infected persons. Funding of this proposal will allow continuation of these studies on the cellular immune response to HCV and further development of the research skills of the principal investigator. This project will be undertaken in the setting of a comprehensive virology research effort involving HIV, herpes simplex virus, Epstein-Barr virus and cytomegalovirus; with numerous opportunities for interaction with other researchers in virology and immunology.